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J. Nat. Prod. 1999, 62, 364-366
Oumarone, Bissaone, and Aissatone, Unusual Prenylated Polyketides from Harrisonia abyssinica Aliou M. Balde´,† Luc Pieters,*,‡ Sandra Apers,‡ Tess De Bruyne,‡ Hilde Van den Heuvel,‡ Magda Claeys,‡ and Arnold Vlietinck‡ Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium, and De´ partement de Pharmacie, Faculte´ de Me´ decine-Pharmacie-Odontostomatologie, Universite´ de Conakry, Conakry, Guinea Received September 2, 1998
Fractionation of the n-hexane extract of the leaves of Harrisonia abyssinica, collected in Guinea, afforded three novel and unusual prenylated polyketides, which were named oumarone (1), bissaone (2), and aissatone (3). Their structures were elucidated by spectroscopic methods, mainly 1D and 2D NMR spectroscopy. Harrisonia abyssinica Oliv. (Simaroubaceae) is a small tree or shrub used widely in African traditional medicine. Extracts of the bark and the root of this plant have been shown to exhibit in vitro antiviral, antibacterial, antifungal, and molluscicidal activities.1 Chemical investigations have led to the identification of various steroids, limonoids, and chromones.2 In this paper we report the isolation and structure elucidation of three novel and unusual prenylated polyketides, which were named oumarone (1), bissaone (2), and aissatone (3). Compounds 1-3 were obtained from the n-hexane extract of powdered stem bark (1 kg) of H. abyssinica, collected in Se´re´dou, Guinea. The EIMS of compound 1 showed a molecular ion at m/z 236, while HREIMS indicated a molecular formula of C15H24O2 (m/z 236.177). 13C NMR signals at δ 196.38, 191.75, and 100.00 were in agreement with a β-diketone in its enolic form (1a). A series of five 13C NMR signals with approximately double intensity could be assigned to two equivalent isoprenyl groups. The remaining CH and CH3 groups (δ 49.05 and 25.15 in 13C NMR) were correlated in a HSQC experiment with 1H NMR signals at δ 2.17 (m) and δ 2.05 (s), respectively. 1H1H COSY and 1H-13C HMBC experiments allowed unequivocal establishment of the structure of 1 as shown. In addition, the presence of a series of minor signals in the 1H and 13C NMR spectra, both for the isoprenyl and the diketone part of the molecule, demonstrated that about 20% of the compound was present as the β-diketone 1b (two carbonyls at δ 207.74 and 202.10, with an isolated CH2 in between at δ 58.38 in the 13C NMR spectrum; the latter corresponded to a two-proton singlet at δ 3.53 in the 1H NMR spectrum). 1H and 13C NMR assignments as well as long-range 1H-13C correlations for 1a and 1b are listed in Table 1. EIMS of 1 revealed diagnostic fragment peaks at m/z 167 (C10H15O2), 151 (C11H19) (loss of 85 amu), 85 (C4H5O2), and 69 (C5H9), which are rationalized in Figure 1. Compound 1, or 8-methyl-5-(3-methyl-2-butenyl)-7-nonene-2,4-dione, for which the trivial name oumarone was adopted, has not been reported before. The EIMS of compound 2 showed a molecular ion at m/z 304, while HREIMS indicated a molecular formula of C20H32O2 (m/z 304.239). From the 1H and 13C NMR data it was evident that the same β-diketone moiety as in 1 was * To whom correspondence should be addressed. Tel. fax, ++32-38202709. E-mail
[email protected]. † Universite ´ de Conakry, Guinea (present address). ‡ University of Antwerp, Belgium.
10.1021/np980379r CCC: $18.00
present, also predominantly in its enolic form (2a). The remaining C15 part could be assigned to three isoprenyl groups; however, no symmetry (as in 1) was observed. Whereas for three isoprenyl groups six methyls were expected (apart from C-1), only five appeared to be present. In contrast, an additional CH2 group occurred at δ 39.79 in the 13C NMR spectrum. This indicated a head-to-tail condensation of two isoprenyl units. Indeed, the 13C NMR assignments for C-6 to C-15 were in agreement with those reported for, for example, the sesquiterpene nerolidol.3 The chemical shift of the C-14 methyl group was indicative of a trans (or E) stereochemistry of the 7,8 double bond. 1H1H COSY and 1H-13C HMBC experiments allowed the unequivocal establishment of the structure of 2 as shown. As for 1, the presence of a series of minor signals in 1H and 13C NMR spectra demonstrated that about 15% of the compound was present as the β-diketone 2b. 1H and 13C NMR assignments as well as long-range 1H-13C correlations for 2a and 2b are listed in Table 2. The EIMS of 2 showed characteristic fragment peaks at m/z 235 (C15H23O2), 167 (C10H15O2), 85 (C4H5O2), and 69 (C5H9), which are rationalized in Figure 1. Compound 2, or 8,12-dimethyl5-(3-methyl-2-butenyl)-7,11-tridecadiene-2,4-dione, for which the trivial name bissaone was adopted, has not been reported before. The EIMS of compound 3 showed a molecular ion at m/z 330, while HREIMS indicated a molecular formula of C20H26O4 (m/z 330.182). In the 13C NMR spectrum four carbonyl signals were present. The 1H-1H COSY spectrum revealed four spin systems: an isoprenyl group, a (CH3)2CH-CH2-CH2 moiety (saturated isoprenyl), an isolated methyl group, and a CH2-CH moiety. The presence of only one quaternary unsaturated carbon signal (at δ 113.43), apart from the double bond in the isoprenyl group and the carbonyl functionalities, was in agreement with the presence of a β-diketone in its enolic form, substituted between the two carbonyls. The corresponding CH group in 1 and 2 occurred around δ 100. Detailed analysis of the longrange 1H-13C correlations allowed construction of a partial structure linking together the four fragments mentioned above. Analysis of the long-range correlations observed for the four carbonyl groups, shown in Figure 2, led unequivocally to structure 3. 1H and 13C NMR assignments, as well as all HMBC correlations observed, are listed in Table 3. The carbonyl group at C-6 showed a typical chemical shift for an isolated ketone, whereas the three other carbonyls were shifted upfield due to enolization.
© 1999 American Chemical Society and American Society of Pharmacognosy Published on Web 01/27/1999
Notes
Journal of Natural Products, 1999, Vol. 62, No. 2 365
Table 1. NMR Spectral Data for 1a Tautomer 1a position 1 2 3 4 5 1′ 2′ 3′ 4′ 5′ a
1H
δ, mult.
2.05, s 5.45, s 2.17, m a: 2.19, m b: 2.26, m 5.05, m 1.67, s 1.59, s
13C
δ
Tautomer 1b 1H
HMBC (C f H)
25.15 191.75 100.00 196.38 49.04 30.61
3 1, 3 1 3, 5, 1′ 2′, 4′, 5′ 2′
121.51 133.45 25.76 17.79
1′, 4′, 5′ 1′, 4′, 5′ 2′, 5′ 2′, 4′
δ, mult.
2.05, s 3.53, s 2.62, m 2.15 2.30, m 5.06, m 1.67, s 1.59, s
13C
δ
HMBC (C f H)
25.15 207.74 58.38 202.10 53.24 29.68
5
120.99 134.15 25.76 17.79
5, 4′, 5′ 1′, 4′, 5′ 2′, 5′ 2′, 4′
1, 3 3, 1′
Recorded in CDCl3 at 400 MHz. Table 2. NMR Spectral Data for 2a Tautomer 2ab position 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 1′
Figure 1. Structure and MS fragmentation (m/z) of 1 and 2 (EIMS).
EIMS of 3 showed a rather complex fragmentation pattern. The high mass range showed fragment ions corresponding to the loss of CH3‚, H2O, CO, and CH3CO‚, at m/z 315, 312, 302, and 287, respectively. The elimination of H2O was in agreement with the presence of keto groups, whereas the loss of CH3CO‚ supported the presence of an acetyl substituent. This was also indicated by the formation of CH3CO+ ions at m/z 43. In the low mass range, the formation of a C5H9+ fragment is energetically favored because the isopentenyl group is attached to a tertiary carbon center and because of resonance stabilization of the C5H9+ ions. In contrast to EIMS, negative-ion FABMS showed more specific fragmentation and only revealed two fragment ions at m/z 219 and 83. A product ion spectrum of [M - H]- ions, obtained using collision-induced dissociation with argon gas at low kinetic energy (30 eV), showed diagnostic product ions at m/z 287, 219, and 83, corresponding to the loss of elements of ketene (CH2dCdO), the combined loss of elements of ketene and C4H8, and the formation of C4H3O2- ions. The absolute stereochemistry of 3 has not been determined. The C-6 carbonyl bridge between C-5 and C-7 only allows a (5R,7S)- or a (5S,7R)-configuration. In the case of a (5R,7S)-stereochemistry (as shown for 3), a (2′S)-configuration was preferred because of high ring strains observed for the (2′R)-isomer when constructing a molecular model. In conclusion, the absolute stereochemistry of 3 is (5R,7S,2′S) or (5S,7R,2′R). The IUPAC name for compound 3, for which the trivial name aissatone was adopted, with the stereochemistry as shown in Figure 2 but in its tetraketo-form, is (1S,5R,7R)-9-acetyl-4,4-di-
2′ 3′ 4′ 5′
1H
δ, mult.
2.05, s 5.45, s 2.18, m a: 2.18, m b: 2.25, m 5.05, m 1.96, m 2.04, m 5.05, m 1.67, s 1.58, s 1.58, s a: 2.18, m b: 2.25, m 5.05, m 1.67, s 1.58, s
13C
δ
25.17 191.78 100.04 196.31 49.04 30.54 121.50c 137.11 39.79 26.72 121.58c,d 133.41e 25.68f 16.10 17.67g 30.54 124.24d 131.39e 25.76f 17.79g
HMBC (CfH) 1, 3, 5 1 3 6, 1′ 9, 14 9, 14 10, 14 9 9, 13, 15 13, 15 15 13 4′, 5′ 4′, 5′ 5′ 4′
a Recorded in CDCl at 400 MHz. b Tautomer 2b: 1H NMR δ 3 3.53 (s, H-3), 2.61 (m, H-5); 13C NMR δ 207.72 (C-2), 58.36 (C-3), 202.06 (C-4), 53.24 (C-5), 120.82, 120.98, 124.11 (C-7, C-11, C-2′), 39.76 (C-9), 26.01 (C-10); HMBC correlations observed between C-2 and H-3, C-4 and H-3, C-4 and H-5. Other signals and HMBC correlations are identical as in 2a, or not observed because of excessive overlap. c-g Assignments bearing the same superscript may be interchanged.
Figure 2. Structure and HMBC correlations involving carbonyl groups observed for 3 (C f H).
methyl-7-(3-methyl-2-butenyl)-tricyclo[5.3.1.01,5]undecane8,10,11-trione. This skeleton has not been reported before from nature. Biogenetically, the C-1 through C-8 chain in 1-3 can be considered as a polyketide consisting of four C2 units (one acetate and three malonates), substituted at C-5 with two isoprenyl chains, one of which is saturated (C-1′ through C-5′). The C-6′-C-7′ moiety is presumed to be an additional C2 component originating from malonyl-CoA. This biogenetic hypothesis was supported by the presence of 1 and
366 Journal of Natural Products, 1999, Vol. 62, No. 2 Table 3. NMR Spectral Data for 3a position 1 2 3 4 5 6 7 8 9 10 11 12 13 1′ 2′ 3′ 4′ 5′ 6′ 7′ a
1H
δ, mult., J
2.36, s
a: 2.33, m b: 2.69, dd, 14.8, 9.4 5.25, m 1.70, s 1.72, s a: 1.85, m b: 1.90, m 2.18, dd, 10.1, 5.4 0.98, s 0.73, s a: 1.59, m b: 1.64, m a: 1.88, m b: 2.44, ddd, 13.2, 8.3, 2.5
13C
δ
32.07 201.67 113.43 197.11 70.18 213.85 76.17 194.47 27.16
HMBC (C f H) 1 9b, 1′ 9, 12, 13, 1′ 9b, 1′, 2′, 7′a 2′, 6′, 7′ 2′, 7′ 1′b
122.34 9, 12, 13 134.32 9, 12, 13 26.05 10, 13 17.96 10, 12 29.55 9, 2′ 57.32 43.11 28.41 22.29 44.62
1′, 4′, 5′, 6′, 7′b 1′, 2′, 4′, 5′, 6′, 7′b 2′, 5′, 6′ 2′, 4′, 6′ 4′, 5′, 7′
21.69
6′
Recorded in CD3OD at 600 MHz.
2, containing three condensed C2 units (one carbon being removed by decarboxylation), substituted at C-5 with two isoprenyl units or one isoprenyl and one geranyl unit. Alternatively, in 3 the C-7′-C-6′-C-3′-C-4′-C-5′ moiety can be considered as an isoprenyl group as well, with C-1′C-2′ being the additional C2 unit. Biogenetically, 3 shows some similarity with humulone and related hop and beer bitter acids, which are prenylated acylphloroglucinols. Phloroglucinol (or 3,5-dihydroxyphenol) consists of three condensed C2 units.4 The basic acylphloroglucinol structure is still present in 3, but, because of the quaternization of C-5 and C-7′, the aromaticity is lost. Experimental Section General Experimental Procedures. EIMS, including high-resolution measurements (HREIMS) and FABMS, was performed on a VG70SEQ instrument (Micromass, Manchester, UK), using glycerol as the liquid matrix. 1D 1H and 13C NMR spectra (including APT and DEPT), and 2D 1H-1H COSY, 1H-13C HMQC or HSQC (one-bond correlations), and 1 H-13C HMBC (long-range correlations) spectra were recorded on Bruker DRX-400 (400 MHz) and Bruker AM-600 (600 MHz) instruments. UV spectra were recorded on a Uvikon 931 instrument (Kontron). Optical rotations were measured on a Perkin-Elmer model 241 polarimeter. Plant Material. The roots and stem bark of Harrisonia abyssinica Oliv. (Simaroubaceae) were collected around Se´re´dou (Guinea) in July 1987. The plant was identified at the Department of Botany of the Research Center of Medicinal Plants, Se´re´dou (Guinea), where a voucher specimen is kept. Extraction and Isolation. All compounds were obtained from the n-hexane extract of powdered stem bark (1 kg) of H.
Notes
abyssinica. After evaporation to dryness under reduced pressure, the organic extract was subjected repeatedly to column chromatography on Si gel eluted with n-hexane and a n-hexane-CHCl3 gradient. The most lipophilic fraction was purified by a combination of column chromatography on Si gel using the same solvent system and preparative TLC on Si gel, using the upper layer of cyclohexane-MeCN-EtOAc-HCO2H (80: 80:30:1) as the mobile phase. In this way, three lipophilic subfractions (I-III) were obtained, subfraction I being the most lipophilic. Further purification by repetitive preparative TLC on silanized Si gel plates using MeOH-H2O (1:1 and 8:2) as the mobile phase yielded compounds 1 (24 mg) and 2 (49 mg) from subfraction II and compound 3 (12 mg) from subfraction III. Structures were elucidated by MS and NMR spectroscopy. Oumarone [8-methyl-5-(3-methyl-2-butenyl)-7-nonene2,4-dione] (1): obtained as a yellowish oil; UV (CHCl3) λmax 278 nm; 1H NMR (CDCl3, 400 MHz), see Table 1; 13C NMR (CDCl3, 100 MHz), see Table 2; EIMS m/z 236 [M]+ (13), 167 (100), 151 (38), 85 (98), 69 (84); HREIMS m/z 236.177 (calcd for C15H24O2, 236.178). Bissaone [8,12-dimethyl-5-(3-methyl-2-butenyl)-7,11tridecadiene-2,4-dione] (2): obtained as a yellowish oil; [R]35D -2.5° (c 0.20, CHCl3); UV (CHCl3) λmax 278 nm; 1H NMR (CDCl3, 400 MHz), see Table 2; 13C NMR (CDCl3, 100 MHz), see Table 2; EIMS m/z 304 [M]+ (8), 235 (22), 167 (50), 85 (87), 69 (100); HREIMS m/z 304.239 (calcd for C20H32O2, 304.238). Aissatone [(1S,5R,7R)-9-acetyl-4,4-dimethyl-7-(3-methyl-2-butenyl)-tricyclo [5.3.1.01,5]undecane-8,10,11-trione] (3): obtained as an amorphous powder; [R]35D -7.3° (c 0.24, MeOH); UV (MeOH) λmax 267 nm; 1H NMR (CD3OD, 600 MHz), see Table 3; 13C NMR (CD3OD, 150 MHz), see Table 3; EIMS m/z 330 [M]+ (100), 315 (52), 312 (23), 302 (26), 287 (31); FABMS (neg. ion mode) m/z 329 [M - H]-, 219, 83; HREIMS m/z 330.182 (calcd for C20H26O4, 330.182). Acknowledgment. S.A., T.D.B., and M.C. are researchers associated with the Fund for Scientific Research (FWOFlanders, Belgium). The FWO is also acknowledged for financial support (grant no. G.0119.96). A.B. was a recipient of a grant of the Algemeen Bestuur voor Ontwikkelingssamenwerking (ABOS, Belgium). V. Wray (GBF, Braunschweig, Germany) is kindly acknowledged for recording NMR spectra at 600 MHz. References and Notes (1) Balde´, A. M.; Pieters, L.; De Bruyne, T.; Geerts, S.; Vanden Berghe, D.; Vlietinck, A. Phytomedicine 1995, 1, 299-302. (2) (a) Kubo, I.; Tanis, S. P.; Lee, Y. W.; Miura, I.; Nakanishi, K.; Chapya, A. Heterocycles 1976, 5, 485-498. (b) Nakanishi, K. J. Nat. Prod. 1982, 45, 15-26. (c) Okorie, D. A. Phytochemistry 1982, 21, 24242426. (d) Hassanali, A.; Bentley, D. M.; Slavin, A. M. Z.; Williams, D. J.; Shepard, R. N.; Chapya, A. W. Phytochemistry 1987, 24, 573575. (e) Balde´, A. M.; Vanhaelen, M.; Daloze, D. Phytochemistry 1988, 24, 942-943. (f) Balde´, A. M. Biological Investigations on Three Medicinal Plants Widely Used in Guinean Traditional Medicine. Ph.D. Thesis, University of Antwerp, Belgium, 1990; pp 327-373. (g) Rajab, M. S.; Rugutt, J. K.; Fronczek, F. R.; Fischer, N. H. J. Nat. Prod. 1997, 60, 822-825. (3) Wenkert, E.; Buckwalter, B. L.; Burfitt, I. R.; Gasic, M. J.; Gottlieb, H. E.; Hagaman, E. W.; Schell, F. M.; Wovkulich, P. M. In Topics in Carbon-13 NMR Spectroscopy; Levy, G. C., Ed.; Wiley: New York, 1976; Vol. 2, Chapter 2, pp 81-121. (4) Teuscher, E. Biogene Arzneimittel. Wissenschaftlige Verlagsgesellschaft mbH: Stuttgart, 1997; pp 182-185.
NP980379R
